慢病毒包装

 

产品简介

 

       慢病毒(Lentivirus)载体是以 HIV-1 (人类免疫缺陷1型病毒)为基础发展起来的基因治疗载体。具有感染谱广泛、可以有效感染分裂期和静止期细胞、长期稳定表达外源基因等优点,因此成为导入外源基因的有力工具。现在慢病毒系统已经被广泛应用到各种细胞系的基因过表达、RNA干扰、microRNA研究以及活体动物实验中。

 

汉恒服务项目

 

      汉恒生物科技(上海)有限公司提供慢病毒载体构建、RNAi(shRNA)、miRNA、过表达和干扰慢病毒包装以及基于慢病毒技术的稳定细胞系构建服务。详情可查询:病毒现货库

   

 

整体实验流程

 

(一) 实验流程(1、2、3为并列步骤)

 

      1. 慢病毒过表达质粒载体的构建 设计上下游特异性扩增引物,同时引入酶切位点,采用PCR技术(采用高保真KOD酶,3K内突变率为0%)从模板中(cDNA质粒或者文库)调取目的基因CDS区(coding sequence)连入T载体。将CDS区从T载体上切下,装入慢病毒过表达质粒载体。

 

     2. 慢病毒干扰质粒载体的构建 合成siRNA对应的DNA颈环结构,退火后连入慢病毒干扰质粒载体。

 

     3. miRNA载体构建从 Genomic 中调取基因相应的Mir 前体, 并在调取引物中引入酶切位点。

 

     4. 慢病毒载体的包装与浓缩纯化 制备慢病毒穿梭质粒及其辅助包装原件载体质粒(两质粒包装系统),三种质粒载体分别进行高纯度无内毒素抽提,共转染293T细胞,转染后6 h 更换为完全培养基,培养48和72h后,分别收集富含慢病毒颗粒的细胞上清液,病毒上清液通过超离心浓缩病毒。

 

(二)流程图

 

 

 

提供产品

 

 

 参考文献

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   3、miR-451a Inhibited Cell Proliferation and Enhanced Tamoxifen Sensitive in Breast Cancer via Macrophage Migration Inhibitory Factor;
     (2015年发表于BioMed Research International,作者:李红)
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     (2014年发表于Tumor Biology,作者:Zhenlin Yang)
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     (2014年发表于Cell Physiol Biochem,作者:Ke Zhou, and Biao Cheng)
   8、MicroRNA-23a is involved in tumor necrosis factor-α induced apoptosis in mesenchymal stem cells and myocardial infarction;
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     (2014年发表于Toxicology and Applied Pharmacology)
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     (2015年发表于Cell Physiol Biochem,作者:Jie Chao)
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     (2015年发表于Tumor Biology,作者:Jianru Xiao)
   14、CRCT1 regulated by microRNA-520 g inhibits proliferation and induces apoptosis in esophageal squamous cell cancer;
     (2015年发表于Tumor Biology,作者:Zhiming Chen)
   15、Overexpression of Hiwi Inhibits the Growth and Migration of Chronic Myeloid Leukemia Cells;
     (2015年发表于Cell Biochemistry and Biophysics,作者:Ziling Liu)
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     (2015年发表于Molecular Immunology,作者:Tao Jianga)
   18、MiR-146b-5p promotes metastasis and induces epithelial-mesenchymal transition in thyroid cancer by targeting ZNRF3;
     (2015年发表于Cell Physiol Biochem,作者:Xiaoping Zhang and Youben Fan)
   19、HSF1 Relieves Amyloid-β-Induced Cardiomyocytes Apoptosis;
     (2015年发表于Cell Biochemistry and Biophysics,作者:Beiru Zhang)
   20、Interaction between TNFR1 and TNFR2 dominates the clinicopathologic features of human hypopharyneal carcinoma;
     (2015年发表于Tumor Biology,作者:Qi Song)
   21、Periostin mediates TGF-β-Induced epithelial mesenchymal Transition in prostate cancer cells;
     (2015年发表于Cell Physiol Biochem,作者:Chuanyu Sun and Guowei Xia)
   22、CD49f acts as an inflammation sensor to regulate differentiation, adhesion, and migration of human mesenchymal stem cells;
     (2015年发表于STEM CELLS,作者:Ran Xiao)
   23、MCPIP1 regulates alveolar macrophage apoptosis and pulmonary fibroblast activation after in vitro exposure to silica;
     (2015年发表于Journal of Neuroinflammation ,作者:姚红红)
   24、MicroRNA-33b inhibits tumor cell growth and is associated with prognosis in colorectal cancer patients;
     (2015年发表于Clinical and Translational Oncology,作者:孙荣勋)
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     (2015年发表于ToxSci Advance Access,作者:Jie Chao)
   26、miR‐152 induces human dental pulp stem cell senescence by inhibiting SIRT7 expression;
     (2015年发表于FEBS letters,作者:Jingping Liang)
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     (2015年发表于International Journal of Molecular Medicine,作者:Hui Li)
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     (2015年发表于Biomedicine & Pharmacotherapy,作者:Jianqing Tian )
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     (2016年发表于Life Sciences,作者:Li-Mimg Zhou、Yuan-Yuan Zhang )
   30、MicroRNA-30a promotes chondrogenic differentiation of mesenchymal stem cells through inhibiting Delta-like 4 expression;
     (2016年发表于Life Sciences,作者:Ye Tian)
   31、MicroRNA-33 promotes the replicative senescence of mouse embryonic fibroblasts by suppressing CDK6;
     (2016年发表于Biochemical and Biophysical Research Communications,作者:Xinguang Liu )
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     (2015年发表于PloS one,作者:Tao Meng)
   33、Repair of critical-sized bone defects with anti-miR-31-expressing bone marrow stromal stem cells and poly (glycerol sebacate) scaffolds;
     (2014年发表于European cells & materials,作者:Deng Yuan)
   34、MicroRNA-22 induces endothelial progenitor cell senescence by targeting AKT3;
     (2014年发表于Cell Physiol Biochem ,作者:Xu Z)
   35、A lentiviral sponge for miRNA-21 diminishes aerobic glycolysis in bladder cancer T24 cells via the PTEN/PI3K/AKT/mTOR axis;
     (2015年发表于Tumor Biology,作者:Changjun Yin)
   36、p53/PUMA expression in human pulmonary fibroblasts mediates cell activation and migration in silicosis;
     (2015年发表于Sci Rep,作者:Jie Chao)
   37、Endocytosis mediated by Caveolin-1 inhibits activity of matrix metalloproteinase-2 in human renal proximal tubular cells under hypoxia;
     (2016年发表于Int J Clin Exp Pathol,作者: Jun Dou )
   38、MTSS1 suppresses cell migration and invasion by targeting CTTN in glioblastoma;
     (2015年发表于Journal of Neuro-Oncology,作者:Qigui Qi)
   39、PFDN1, an indicator for colorectal cancer prognosis, enhances tumor cell proliferation and motility through cytoskeletal reorganization;
     (2015年发表于Medical Oncology,作者:Minhua Zheng )
   40、Human Dental Pulp Stem Cells via the NF-κB Pathway;
     (2015年发表于Cell Physiol Biochem,作者:Jingping Liang )
   41、Clinical and therapeutic significance of sirtuin-4 expression in colorectal cancer;
     (2016年发表于Oncol Rep,作者:Peng Z)
   42、58-F, a flavanone from Ophiopogon japonicus, prevents hepatocyte death by decreasing lysosomal membrane permeability;
     (2016年发表于Sci Rep,作者:Xiaoling Wang)
   43、Study of Arsenic Sulfide in Solid Tumor Cells Reveals Regulation of Nuclear Factors of Activated T-cells by PML and p53;
     (发表于Sci Rep,作者:Chen S1)

 

 
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